Malaria, a disease caused by parasitic protozoans in the genus Plasmodium, is transmitted by mosquitoes to humans. Each year, hundreds of millions of people are infected with this organism and 2-3 million fatalities result, mostly in young children. The problem is most severe in Africa, and is now worsening because of the increasing resistance of parasites to the inexpensive drugs used for prevention and control in the past. A second important strategy for control, reducing mosquito populations through insecticide use, is also threatened because of insecticide resistance, increasing costs and loss of trained personnel. A new target for research has been the interaction between the mosquito and the parasite. The long-term goal of the research presented here is to understand the molecular basis for incompatibility between mosquitoes and malaria parasites because this may suggest ways to enhance mechanisms that cause incompatibility through genetic or chemical manipulation of vector mosquitoes. The proposed research will focus on characterizing proteins and genes that are involved in the mosquito's humoral immune responses, which can lead to the death of parasites. In particular, serine proteases are important enzymes in generation of two parasite-killing mechanisms, melanotic encapsulation and the antimicrobial response. In the African malaria vector, Anopheles gambiae, three serine protease genes, AgSp14D1, AgSp14D2, and AgSp14A, show changes in transcript levels after bacterial injections or infection with malaria parasites. These molecules will be characterized and tested for involvement in immune responses against parasites in vitro and in vivo. In addition, prophenoloxidase activating serine proteases, which are important in triggering parasite melanization, will be biochemically purified and then cloned and characterized. Finally, several A. gambiae serine proteases that exhibit domain structures similar to other enzymes involved in immune responses will be characterized at the mRNA and protein levels.